Proteases participate in a great variety of biological processes, from the digestion of proteins in the diet in the lumen of the gastrointestinal tract to controlling the cell cycle. An important role of proteases is their participation in signal transduction processes, whether this is by proteolytically cutting ligands or cell surface receptors to generate active ligands, or, on the contrary, to degrade and inactivate agonists of certain receptors. One type of these cell surface receptors which are attacked by the proteases generating active ligands is the family of protease-activated G protein-coupled receptors, PAR (proteinase-activated receptors), for which a protease cuts in a specific place on the extracellular N-terminal domain of the receptor, and this cut means that a new N-terminal domain is exposed which acts as an anchored ligand, bringing about the beginning of the signal transduction [Hollenberg M. D., “Physiology and pathophisiology of Proteinase Activated Receptors (PARs): proteinases as hormone-like signal messengers: PARs and more”, (2005), J. Pharmacol. Sci., 97, 8-13]. The family of PARs has four members, PAR-1 to PAR-4, which are activated by a large number of proteases, such as coagulation cascade proteases such as thrombin or the TF-FVIIa-FXa complex, inflammatory cell proteases such as mast cell proteases (tryptase) and leucocytes (cathepsin G, elastase and proteinase-3), digestive tract proteases such as trypsins and pancreatic and extrapancreatic trypsinogens, tissue proteases such as kallikreins, as well as non-mammalian proteases, such as bacterial proteases, fungi, mites or insects [Ossovskaya V. S. and Bunnet N. W., “Protease Activated Receptors-contribution to physiology and disease”, (2004), Physiol. Rev., 84, 579-621]. There is a certain selectivity in PAR activation; thrombin, for example, can activate PAR-1, PAR-3 and PAR-4 with varying degrees of intensity, but does not activate PAR-2, whilst trypsin and tryptase or mast cell proteases activate PAR-2.
Specifically, PAR-2 is widely distributed in the human body, including the skin, the gastrointestinal tract and the circulatory, respiratory and nervous systems, modulating different physiological functions such as coagulation, proliferation and survival, inflammation, neurotransmission and pain. In pathological conditions, such as hemostasis or inflammation, both an overexpression of PAR-2 has been observed [Nystedt S. et al. “The Proteinase-activated Receptor 2 is induced by inflammatory mediators in human endothelial cells. Comparison with the thrombin receptor”, (1996), J. Biol. Chem., 1271(25), 14910-14915] as well as an overproduction of proteases capable of activating PAR-2.
In the skin, PAR-2 is expressed abundantly in almost all cells types, especially in keratinocytes, and is particularly important in the stratum granulosum, which means its expression can depend on the state of epidermal differentiation. In the stratum corneum there are three families of proteases; specific epidermal serine proteases such as kallikrein-5 (SCTE or stratum corneum tryptic enzyme) or kallikrein-7 (SCCE or stratum corneum chymotryptic enzyme), cysteine proteases such as cathepsins C, L, and V (stratum corneum thiol proteases), and at least one aspartate protease (cathepsin D). The activity of these proteases is closely regulated by specific inhibitors and acts as a mediator of several cells responses in the skin, such as inflammation and immune responses, chemotaxis, cytokine expression, vascular function, tissue and apoptosis repair. As well as said endogenous proteases, signaling in the epidermis can also be due to some exogenous allergen proteases such as domestic mites, cockroaches, pollen, certain bacteria and fungi. PAR-2 is a sensor for all these proteases, playing an important role in the maintenance of homeostasis of the barrier function of the skin. The participation of PAR-2 in homeostasis of the barrier function of the skin has been proven both in cell cultures and in animal models; the activation of PAR-2 in keratinocytes in culture inhibits cell proliferation, which is consistent with the delay in recovery of the barrier and inhibition of the secretion of lamellar bodies observed after the topical application of a PAR-2 peptide agonist or allergens on the skin of mice, whilst an increase in the secretion of lamellar bodies and an accelerated recovery of the barrier function after the disruption to said barrier function has been observed in PAR-2 knockout mice, in comparison with their wild phenotype littermates [Derian C. K. et al., “Differential regulation of human keratinocyte growth and differentiation by a novel family of protease-activated receptors”, (1997), Cell Growth Differ., 8(7), 743-749; Hachem J.-P. et al., “Serine protease signaling of epidermal permeability barrier homeostasis”, (2006), J. Invest. Dermatol., 126(9), 2074-2086; Jeong S. K. et al., “Mite and cockroach allergens activate Protease-Activated Receptor 2 and delay epidermal permeability barrier recovery”, (2008), J. Invest. Dermatol., 128(8), 1930-1939].
The abnormal expression or abnormal activity of proteases and, therefore, the overactivation of PAR-2 is associated with skin disorders or diseases such as atopic dermatitis, Netherton's syndrome, psoriasis and peeling skin syndrome [Komatsu N. et al., “Elevated human tissue kallikrein levels in the stratum corneum and serum of peeling skin syndrome-type B patients suggests an over-desquamation of corneocytes”, (2006), J. Invest. Dermatol., 126(10), 2338-2342]. Elevated levels of PAR-2 have also been described in skin inflammatory or immune diseases such as lichen planus, atopic dermatitis, psoriasis [Carvalho R. F. et al. “Increased mast cell expression of PAR-2 in skin inflammatory diseases and release of IL-8 upon PAR-2 activation”; (2010), Exp. Dermatol., 19(2), 117-122] or rosacea [Hachem J.-P. et al., “Serine protease signaling of epidermal permeability barrier homeostasis”, (2006), J. Invest. Dermatol., 126(9), 2074-2086].
PAR-2 is also involved in the development of inflammatory processes. Many of the cells which orchestrate the response of the immune system during inflammation express PAR-type receptors; for example eosinophilic infiltrates express PAR-2 [Miike S. et al., “Trypsin induces activation and inflammatory mediator release from human eosinophils through Protease-Activated Receptor-2”, (2001), J. Immunol., 167(11), 6615-6622]. During the inflammatory processes potential endogenous PAR-2 activators are released, such as leukocyte elastase, mast cell tryptase, proteinases of the trypsin family produced by keratinocytes and proteases of the fibrinolytic cascade such as factors FVIIa or FXa. These components activate PAR-2 in keratinocytes, endothelial cells, inflammatory cells and dermal sensory nerves to amplify inflammation by over-regulating inflammatory mediators. The activation of PAR-2 causes nitric oxide-dependent vasodilation [Saifeddine M. et al., “Rat Proteinase-Activated Receptor-2 (PAR-2): cDNA sequence and activity of receptor-derived peptides in gastric and vascular tissue”, (1996), Br. J. Pharmacol., 118, 521-530; Sobey C. G. et al., “Activation of Protease-Activated Receptor-2 (PAR-2) elicits nitric oxide-dependent dilatation of the basilar artery in vivo”, (1998), Stroke, 29(7), 1439-1444], induces extravasation of plasma proteins and infiltration of neutrophils [Vergnolle N. et al., “Characterization of the inflammatory response to Proteinase-Activated Receptor-2-activating peptides in the rat paw”, (1999), Br. J. Pharmacol., 127(5), 1083-1090; Kawabata A. et al., “Increased vascular permeability by a specific agonist of Protease-Activated Receptor-2 in rat hindpaw”, (1998), Br. J. Pharmacol., 125(3), 419-422], and stimulates the secretion of pro-inflammatory cytokines [Hou L. et al., “Immunolocalization of Protease-Activated Receptor-2 in skin: receptor activation stimulates interleukin-8 secretion by keratinocytes in vitro”, (1998), Immunology, 94(3), 356-362]. Furthermore, it has been described that in an animal model of contact hypersensitivity on mice ears, PAR-2 knockout mice show a reduction in the swelling of ears and in the volume of inflammatory infiltrates, corroborating that PAR-2 plays a role as an inflammatory mediator in allergic dermatitis [Kawagoe J. et al., “Effect of Protease-Activated Receptor-2 deficiency on allergic dermatitis in the mouse ear”, (2002), Jpn. J. Pharmacol., 88(1), 77-84]. It is also described that PAR-2 intervenes in the development of oral diseases and disorders such as periodontitis; the protease of the microorganism Porphyromonas gingivalis presents PAR-2 in the active oral cavity and induces the secretion of IL-6, causing the infiltration of granulocytes in the gums and periodontitis through a mechanism which comprises the release of prostaglandin and the activation of matrix metalloproteinases with the consequential destruction of the collagen tissue supporting the teeth [Lourbakos A. et al., “Arginine-specific protease from Porphyromonas gingivalis activates Protease-Activated Receptors on human oral epithelial cells and induces interleukin-6 secretion”, (2001), Infect. Immun., 69(8), 5121-5130; Holzhausen M., Spolidorio L. C. and Vergnolle N., “Role of Protease-Activated Receptor-2 in inflammation, and its possible implications as a putative mediator of periodontitis”, (2005), Mem. Inst. Oswaldo Cruz., 100(1), 177-180].
Chronic inflammatory processes are also mediated by PAR-2, as is the case of rheumatoid arthritis. Activation of PAR-2 in a mouse's knee joint with PAR-2-activating peptides results in a swelling of the joint and hyperemia, clear indicators of inflammation. The duration of the inflammation and swelling can vary according to the nature of the PAR-2-activating peptide. At the same time, an increase in the expression of PAR-2 in the inflamed synovium is detected, as well as in adjacent muscle and skin, presumably by discharge of adjuvant into said tissues during induction, this indicating that the increase in PAR-2 expression is associated with chronic inflammatory responses in different types of tissue. However, mutant PAR-2 knockout mice are protected from arthritis induced by intra-articular and peri-articular injections of CFA [Ferrell W. R. et al., “Essential role for Proteinase-Activated Receptor-2 in arthritis”, (2003), J. Clin. Invest., 111(1), 35-41; Kelso E. B. et al., “Therapeutic promise of Proteinase-Activated Receptor-2 antagonism in joint inflammation”, (2006), J. Pharmacol. Exp. Ther., 316(3), 1017-1024].
PAR-2 does not just participate in nociception, but also in the transmission of pain. The injection of sub-inflammatory doses of PAR-2-agonists in murinae induces hyperalgesia sustained by mechanical and thermal stimuli, whilst this somatic hyperalgesia is absent in PAR-2 knockout animals [Vergnolle N. et al., “Proteinase-Activated Receptor-2 and hyperalgesia: a novel pain pathway”, (2001), Nat. Med., 7(7), 821-826]. The involvement of PAR-2 has also been described, and in particular in inflammatory pain, in visceral pain and pain due to cancer. For example, pancreatitis is associated with premature activation of trypsinogen in the pancreas which induces hyperalgesia by a PAR-2-dependant mechanism [Hoogerwerf W. A. et al., “The Proteinase-Activated Receptor-2 is involved in nociception”, (2001), J. Neurosci., 21(22), 9036-9042] as well as visceral pain in different diseases of the digestive tract such as irritable bowel syndrome, ulcerative colitis and Crohn's disease [Cenac N., “Role for protease activity in visceral pain in irritable bowel syndrome”, (2007), J. Clin. Invest., 117(3), 636-647]. In the same way, mechanical allodynia caused in cancerous processes disappears in PAR-2 knockout mice [Lam D. K. and Schmidt B. L., “Serine proteases and Protease-Activated Receptor-2-dependent allodynia: a novel cancer pain pathway”, (2010), Pain, 149(2), 263-272]. The document WO 2009/117481 A1 describes the use of PAR-2 activity inhibitors for the treatment of chronic pain, inflammatory pain, postoperative pain, neuropathic pain, pain due to fractures, osteoporotic fractures, cancer or joint pain caused by gout, among others.
PAR-2 notably contributes to neurogenic inflammation, as it is expressed in nociceptive peptidergic neurons of the peripheral system, which are responsible for this inflammation. During neurogenic inflammation, different endogenous serine proteases such as mast cell tryptase and keratinocyte trypsin activate PAR-2 in sensory nerve ends to release a calcitonin gene-related peptide (CGRP) and substance P (SP). These neuropeptides are pro-inflammatory: they induce vasodilatation, edema, and leukocyte recruitment, which results in neurogenic inflammation [Steinhoff M. et al. “Agonists of Proteinase Activated Receptor-2 induce inflammation by a neurogenic mechanism”, (2000), Nat. Med., 6(2), 151-158].
PAR-2 activation also mediates in the induction of itching [Shimada S. G., et al., “Scratching behavior in mice induced by the Proteinase-Activated Receptor-2 agonist, SLIGRL-NH2”, (2006), Eur. J. Pharmacol., 530(3), 281-283], and this itching is independent of histamine. PAR-2 activation triggers the release of substance P, which, as well as causing itchiness, promotes the continued activation of mast cells via TRK receptors and the resulting release of tryptase, which in turn activates PAR-2 [Greaves M., “Recent advances in pathophysiology and current management of itch”, (2007), Ann. Acad. Med. Singapore., 36(9), 788-792]. The levels of tryptase are abnormally high in disorders or diseases of which involve itching, such as in atopic dermatitis, in which the concentrations of tryptase are up to four times higher than those observed in healthy skin [Steinhoff M. et al., “Proteinase-Activated Receptor-2 mediates itch: a novel pathway for pruritus in human skin”, (2003), J. Neurosci., 23(15), 6176-6180]. It is described that PAR-2 antagonists are capable of inhibiting trypsin-elicited scratching [Costa R. et al. “Evidence for the role of neurogenic inflammation components in trypsin-elicited scratching behaviour in mice”, (2008), Br. J. Pharmacol., 154(5), 1094-1103]. This property opens the door to the treatment of different conditions, disorders or diseases which involve itching through PAR-2 activity inhibitors, such as dermatitis, including contact dermatitis and atopic dermatitis, urticaria, food allergies or allergies to insect bites, among others.
PAR-2 sensitizes the transient receptor potential vanilloid 1 (TRPV-1), which belongs to the TRP channel superfamily, amplifying the response to pain, inflammation and itching [Amadesi S. et al., “Protease-Activated Receptor 2 Sensitizes the Capsaicin Receptor Transient Receptor Potential Vanilloid Receptor 1 to Induce Hyperalgesia”, (2004), J. Neurosci., 24(18), 4300-4312; Dai Y. et al., “Proteinase-Activated Receptor 2-Mediated Potentiation of Transient Receptor Potential Vanilloid Subfamily 1 Activity Reveals a Mechanism for Proteinase-Induced Inflammatory Pain”, (2004), J. Neurosci., 24(18), 4293-4299].
PAR-2 is also expressed in neurons and astrocytes of the central nervous system in humans and rodents, and has been related to the pathogenesis associated with ischemia and neurodegeneration [Smith-Swintowski V. L. et al., “Protease-Activated Receptor-2 (PAR-2) is present in the rat hippocampus and is associated with neurodegeneration”, (1997), J. Neurochem., 69(5), 1890-1896] as well as with the development of multiple sclerosis and in experimental autoimmune encephalomyelitis (EAE) [Noorbakhsh F. et al., “Proteinase-Activated Receptor-2 modulates neuroinflammation in experimental autoimmune encephalomyelitis and multiple sclerosis”, (2006), J. Exp. Med., 203(2), 425-435].
PAR-2 also plays an important role in regulating pigmentation. The exposure to UV radiation induces an overexposure of PAR-2 in keratinocytes [Scott G. et al., “Protease-Activated Receptor-2, a receptor involved in melanosome transfer, is upregulated in human skin by ultraviolet irradiation”, (2001), J. Invest. Dermatol., 117(6), 1412-1420], whose activation induces melanoma phagocytosis, which involves a transfer of melanocyte melanin to the keratinocyte with the resulting darkening of the epidermis [Seiberg M., “Keratinocyte-melanocyte interactions during melanosome transfer”, (2001), Pigment Cell Res., 14(4), 236-242]. Skin coloration has been a concern for human beings for many years. In particular, the capacity to eliminate hyperpigmentation, whether due to age (marks, freckles or the general aging of the skin), or due to disorders or diseases (melasma, chloasma, post-inflammatory hyperpigmentation) is of interest for individuals who want an even-looking skin complexion. Likewise, when exposure to UV radiation is prolonged or excessive, cancerous hyperpigmented lesions or melanomas can develop [Dooley T. P., “Recent advances in cutaneous melanoma oncogenesis research”, (1994), Onco. Res., 6, 1-9] as well as benign hyperpigmented marks due to photoaging. It is described that PAR-2 inhibition has a depigmenting effect on the skin [Seiberg M. et al., “Inhibition of melanosome transfer results in skin lightening”, (2000), J. Invest. Dermatol., 115(2), 162-167], therefore the treatment of skin with PAR-2 activity inhibitors is a valid strategy to lessen skin pigmentation, as described in documents EP 0948308 B1, EP 1077063 B1, U.S. Pat. No. 6,750,229 B2 and EP 1139974 A1.
However, the use of whitening or depigmenting compounds, whether for the treatment of hyperpigmented areas or areas close to hypopigmented areas, for aesthetic reasons to lighten the natural skin color, its collateral effect is to increase the risk of damage by UV radiation, as they reduce the quantity of melanin produced by melanocytes. Melanin is the skin's natural photoprotector, as it clears, as heat, over 99.9% of UV radiation absorbed [Meredith P. et al., “Radiative relaxation quantum yields for synthetic eumelanin”, (2004), Photochem. Photobiol., 79(2), 211-216]. This means that less than 0.1% of radiation absorbed will be capable of generating free radicals, which cause direct and indirect DNA damage and, therefore, photoaging. The cosmetic and pharmaceutical industries compensate this lack of protection inherent in the use of whiteners or depigmenting agents with the addition of formulations of photoprotective substances or solar filters. Solar filters protect the skin from UVB radiation, which may cause burns, and from UVA radiation, which damages the skin on a more long-term scale by causing accelerated aging or photoaging. However, many of these substances are potentially irritating, sensitizing or toxic, their use being regulated and even limited or prohibited in different countries. Therefore, there is a need to develop whitening or depigmenting compounds with an intrinsic photoprotective effectiveness which enable the use of additional photoprotectors to be reduced.
There is also expression of PAR-2 receptors in respiratory channels, in ciliated and non-ciliated epithelial cells, as well as in glands, smooth muscle, vascular smooth muscle cells and endothelial cells [D'Andrea M. et al., “Characterization of Protease-Activated Receptor-2 immunoreactivity in normal human tissues”, (1998), J. Histochem. Cytochem., 46(2), 157-164]. Said receptors are activated by endogenous proteases such as trypsin produced in the airway epithelial cells [Miki M. et al., “Effect of human airway trypsin-like protease on intracellular free Ca2+concentration in human bronchial epithelial cells”, (2003), J. Med. Invest., 50(1-2), 95-107], or isolated tryptase in human pulmonary mast cells [Berger P. et al., “Tryptase and agonists of PAR-2 induce the proliferation of human airway smooth muscle cells”, (2001), J Appl Physiol., 91(3), 1372-1379], as well as by proteases of different allergens, such as cockroaches, mites such as Dermatophagoides pteronyssinus or mold [Sun G. et al., “Interaction of mite allergens Der P3 and Der P9 with Protease-Activated Receptor-2 expressed by lung epithelial cells”, (2001), J. Immunol., 167(2), 1014-1021; King C. et al., “Dust mite proteolytic allergens induce cytokine release from cultured airway epithelium”, (1998), J Immunol., 161(7), 3645-3651; Page K. et al., “Mucosal sensitization to German cockroach involves Protease-Activated Receptor-2”, (2010), Respir. Res., 11(1), 62; Chiu L. L. et al., “Mold allergen, Pen c13, induced IL-8 expression in human airway epithelial cells by activated Protease-Activated Receptor 1 and 2”, (2007), J. Immunol., 178(8), 5237-5244].
PAR-2 activation correlates with the mast cell infiltration observed in the allergic inflammation of airways; in PAR-2 knockout animal models there is a lesser infiltration of eosinophils in the event of allergic inflammation induced in airways, whilst the mutant which overexpresses PAR-2 presents an increase in this response [Schmidlin F. et al., “Protease-Activated Receptor-2 mediates eosinophil infiltration and hyperreactivity in allergic inflammation of the airway”, (2002), J. Immunol., 169(9), 5315-5321]. Likewise, patients with asthma overexpress PAR-2 en respiratory epithelial cells but not in smooth muscle or alveolar macrophages [Knight D. A. et al., “Protease-Activated Receptors in human airways: upregulation of PAR-2 in respiratory epithelium from patients with asthma”, (2001), J. Allergy Clin. Immunol., 108(5), 797-803]. Thus, the inhibition of PAR-2 activity is a useful strategy for the treatment of inflammatory diseases of airways, such as allergic rhinitis, chronic obstructive pulmonary disease, bronchial hyperreactivity and asthma.
PAR-2 is expressed in different tumor cells and tissues [D'Andrea M. R. et al., “Differential expression of Protease-Activated Receptors-1 and -2 in stromal fibroblasts of normal, benign, and malignant human tissues”, (2001), Am. J. Pathol., 158(6), 2031-2041; Elste A. P. and Petersen I., “Expression of proteinase-activated receptor 1-4 (PAR 1-4) in human cancer”, (2010), J. Mol. Histol., 41(2-3), 89-99], and plays an important role in the invasion and tumor growth in different malignant neoplasms, such as in the stomach, colon, pancreas, lungs, airways, prostate, uterus and breast [Darmoul D. et al., “Initiation of human colon cancer cell proliferation by trypsin acting at Protease-Activated Receptor-2”, (2001), Br. J. Cancer, 85(5), 772-779; Ikeda O. et al., “Signal of Proteinase-Activated Receptor-2 contributes to highly malignant potential of human pancreatic cancer by up-regulation of interleukin-8 release”, 2006, Int. J. Oncol., 28(4), 939-946; Jin E. et al., “Protease Activated Receptor [PAR]-1 and PAR-2 participate in the cell growth of alveolar capillary endothelium in primary lung adenocarcinomas”, (2003), Cancer, 97(3), 703-713; Li Z. et al., “Expression of Protease-Activated Receptor-2 (PAR-2) in patients with nasopharyngeal carcinoma: correlation with clinicopathological features and prognosis”, (2009), Pathol. Res. Pract., 205(8), 542-550; Wilson S. et al., “The membrane-anchored serine protease, TMPRSS2, activates PAR-2 in prostate cancer cells”, (2005), Biochem. J., 388(Pt 3), 967-972; Sánchez-Hernández P. E. et al., “Protease-Activated Receptor-2 (PAR-2) in cervical cancer proliferation”, (2008), Gynecol. Oncol., 108(1), 19-26; Morris D. R. et al., “Protease-Activated Receptor-2 is essential for factor VIIa and Xa-induced signaling, migration, and invasion of breast cancer cells”, (2006), Cancer Res., 66(1), 307-314]. PAR-2 activation gives rise to classic intracellular signals including the induction of an extensive repertoire of proangiogenic factors which facilitate the proliferation and migration of tumor cells; the inhibition of PAR-2 activity is therefore a useful strategy to restrict the growth of tumors and metastasis, as described in document US 2006/0104944 A1.
The cosmetic and pharmaceutical industry has tried on many occasions to find molecules or extracts which inhibit PAR-2, such as those described in documents WO 2006/035936 A1, WO 2006/127379 A2, US 2004/0266687 A1, US 2006/0142203 A1 and US 2006/0183664 A1 among others. However, despite the arsenal of existing compounds and/or extracts, the cosmetic and pharmaceutical sector is still interested in developing alternatives to the compounds known in the prior art for the treatment and/or care of those conditions, disorders and/or diseases that improve or are prevented by the inhibition of PAR-2 activity.